摘要
目的探讨微小RNA-940(miR-940)对迁移和侵袭增强因子1(MIEN1)的作用,对人骨肉瘤细胞MG63的迁移和侵袭等能力的影响。方法根据对MG63(武汉大学中南医院医学科学研究中心)的转染处理,将其分为miR-940模拟物(mimics)组、控制(control)对照组、miR-940抑制剂(inhibitor)组和空白(Blank)对照组。分析武汉大学中南医院骨科通过手术切除的12例骨肉瘤病理组织,采用世界卫生组织(WHO)病理分级标准进行良恶性分级,Ⅰ级5例,Ⅱ级3例,Ⅲ级4例。采用免疫组织化学分析MIEN1与肿瘤恶性程度的相关性。体外培养人骨肉瘤细胞MG63及正常成骨细胞hFOB1.19,实时荧光定量聚合酶链反应检测miR-940及MIEN1表达水平,蛋白质印迹法(Western blot)检验MIEN1蛋白表达。对MG63细胞分组转染miR-940模拟物(mimics)、控制(control)对照、抑制剂(inhibitor)和空白对照,48 h后采用实时荧光定量PCR检验miR-940,Western blot检验MIEN1蛋白表达。划痕实验评估转染后细胞的迁移能力。Transwell侵袭实验评价转染后细胞的侵袭能力。采用两样本t检验,多样本方差分析,Pearson相关性分析。结果MIEN1在不同恶性程度的骨肉瘤中的表达水平,与恶性程度呈正相关(r=0.832,P<0.05)。骨肉瘤细胞MG63比正常成骨细胞的miR-940表达水平更低(4.37±0.61比12.41±1.05),差异有统计学意义(t=11.464,P<0.01),而MIEN1表达水平更高(21.01±1.01比5.49±0.38),差异有统计学意义(t=-24.954,P<0.01),Western blot结果显示miR-940与MIEN1的表达水平呈负相关(r=-0.914,P<0.01)。转染后,模拟物组miR-940水平高于控制对照及空白对照组,而抑制剂组则低于对照组(51.16±4.27比4.19±0.47比4.36±0.29比1.18±0.15),差异有统计学意义(F=372.327,P<0.01),Western blot结果表明MIEN1的蛋白表达水平为模拟物组比对照组表达降低77.89%(t=3.174,P<0.05),而抑制剂组比对照组表达增高141.78%(t=-4.318,P<0.05)。划痕实验证实迁移能力miR-940模拟物组低�
Objective To investigate the effect of microRNA-940(miR-940)on migration and invasion enhancer 1(MIEN1),and its effect on the migration and invasion of human osteosarcoma cell MG63.Methods The source of human osteosarcoma cell line MG63 and osteoblast cell line hFOB1.19 was preserved by the Medical Research Center of Zhongnan Hospital of Wuhan University.The miR-940-related transfection materials and reagents were purchased from Wuhan Venosi Biological Company.According to the transfection treatment of MG63 cells,they were divided into miR-940 mimics(mimics)group,control(control)control group,miR-940 inhibitor(inhibitor)group and blank(blank)control group.The pathological tissues of 12 cases of osteosarcoma resected surgically in the Department of Trauma and Micro-Orthopedics,Zhongnan Hospital of Wuhan University were analyzed.The World Health Organization(WHO)pathological grading standard was used to classify benign and malignant tumors.There were 5 cases of gradeⅠ,3 cases of gradeⅡ,and 4 cases of gradeⅢ.The correlation between MIEN1 and tumor malignancy was analyzed by immunohistochemistry.This study has passed the ethical review and approval of the clinical research project of the Medical Ethics Committee of Zhongnan Hospital of Wuhan University,with the approval document batch number:2021020.需要写吗?MG63 cells and hFOB1.19 cells were cultured in vitro,and the expression levels of miR-940 and MIEN1 were detected by real-time fluorescent quantitative PCR.The expression of MIEN1 protein was detected by Western blotting.MG63 cells were grouped and transfected with miR-940 mimics,control,inhibitor and blank control,and the expression of MIEN1 protein was detected by Western blotting.The migratory ability of the cells after transfection was assessed by scratch assay.Transwell invasion assay was used to evaluate the invasive ability of cells after transfection.Two-sample t test,multi-sample analysis of variance,and pearson correlation analysis were used.Results The expression level of MIEN1 in osteosarcom
作者
郭亚旗
胡祥
漆白文
喻爱喜
Guo Yaqi;Hu Xiang;Qi Baiwen;Yu Aixi(Department of Trauma and Micro-Orthopedics,Zhongnan Hospital of Wuhan University,Wuhan 430071,China)
出处
《中华实验外科杂志》
CAS
北大核心
2022年第3期531-534,共4页
Chinese Journal of Experimental Surgery
关键词
骨肉瘤
微小RNA
迁移
侵袭
Osteosarcoma
MicroRNA
Migration
Invasion
